The protein kinase calcium/calmodulin-dependent kinase II (CaMKII) predominantly consists of the β ± and β 2 isoforms in the brain. Although CaMKIIβ ± functions have been elucidated, the isoform-specific catalytic functions of CaMKIIβ 2 have remained unknown. Using knockdown analyses in primary rat neurons and in the rat cerebellar cortex in vivo, we report that CaMKIIβ 2 operates at the centrosome in a CaMKIIβ ±-independent manner to drive dendrite retraction and pruning. We also find that the targeting protein PCM1 (pericentriolar material 1) localizes CaMKIIβ 2 to the centrosome. Finally, we uncover the E3 ubiquitin ligase Cdc20-APC (cell division cycle 20-anaphase promoting complex) as a centrosomal substrate of CaMKIIβ 2. CaMKIIβ 2 phosphorylates Cdc20 at Ser51, which induces Cdc20 dispersion from the centrosome, thereby inhibiting centrosomal Cdc20-APC activity and triggering the transition from growth to retraction of dendrites. Our findings define a new, isoform-specific function for CaMKIIβ 2 that regulates ubiquitin signaling at the centrosome and thereby orchestrates dendrite patterning, with important implications for neuronal connectivity in the brain.